Fermentation preparation method of salmonella pullorum bacteriophage lyase

A technology of Salmonella phage and Salmonella pullorum, applied in the field of biology, can solve the problems of poor antibacterial effect, poor cracking effect of Salmonella pullorum phage lyase, poor stability, etc., and achieve acid-base resistance Wide range, no loss of enzyme activity, good thermal stability

Pending Publication Date: 2021-11-05
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Through the above analysis, the existing problems and defects of the prior art are: the existing prepared Salmonella pullorum phage lyase has poor cleavage effect, poor antibacterial effect and poor stability.

Method used

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  • Fermentation preparation method of salmonella pullorum bacteriophage lyase
  • Fermentation preparation method of salmonella pullorum bacteriophage lyase
  • Fermentation preparation method of salmonella pullorum bacteriophage lyase

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preparation example Construction

[0042] Such as figure 1As shown, the fermentation preparation method of Salmonella pullorum phage lyase provided by the embodiments of the present invention comprises the following steps:

[0043] S101, culturing Salmonella pullorum and Salmonella phage YSP2, and extracting the Salmonella phage YSP2 genome;

[0044] S102, using the whole genome of bacteriophage YSP2 as a template, using primers LySP2-ZF / LySP2-ZR to perform PCR amplification on the lyase target gene LySP2;

[0045] S103, the construction and identification of the expression vector pPICZ-LySP2, the screening and identification of the recombinant Pichia strain X33-pPICZ-LySP2; the induction fermentation and condition optimization of the recombinant Pichia strain X33-pPICZ-LySP2 at the same time.

[0046] Those of ordinary skill in the industry can also adopt other steps to implement the fermentation preparation method of Salmonella pullorum phage lyase provided by the invention, figure 1 The fermentation prepar...

Embodiment 1

[0071] 1. Materials

[0072] Restriction enzymes Xho I, Xba I, Sca I, DNA Marker 15,000, DNA Marker 5000, DNAMarker 2000, 10×Loading Buffer, rTaq enzymes were purchased from Japan TAKARA Company; Tris, SDS, β-mercaptoethanol, persulfate Ammonium and TEMED were purchased from American Amresco; Zeocin antibiotics were purchased from the United States; protein markers (180, 130, 95, 72, 55, 43, 34, 26, 17, 10) were purchased from Thermo; agarose gel DNA recovery reagents Kits, plasmid mini-extraction kits, PCR product purification and recovery kits, yeast genome DNA extraction kits, and viral DNA / RNA extraction kits were purchased from Beijing Tiangen Biochemical Technology Co., Ltd.

[0073] According to the sequence of LySP2 gene of Salmonella pullorum phage YSP2 lyase, a pair of primers were designed and named as LySP2-ZF and LySP2-ZR, and the restriction endonucleases Xho I and Xba I were inserted at the 5' end and 3' end respectively. Restriction site, used for the cloning ...

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Abstract

The invention belongs to the technical field of biology, and discloses a fermentation preparation method of salmonella pullorum bacteriophage lyase, which comprises: culturing salmonella pullorum and salmonella bacteriophage YSP2, and extracting a salmonella bacteriophage YSP2 genome; carrying out polymerase chain reaction (PCR) amplification on a lyase target gene LySP2 by utilizing a primer LySP2-ZF/LySP2-ZR, by taking a whole genome of the bacteriophage YSP2 as a template; constructing and identifying an expression vector pPICZ-LySP2, and screening and identifying a recombinant pichia pastoris strain X33-pPICZ-LySP2; simultaneously carrying out induced fermentation and condition optimization of the recombinant pichia pastoris X33-pPICZ-LySP2. According to the present invention, the pichia pastoris expression strain X33-pPICZ-LySP2 of a recombinant lyase LySP2 gene is constructed.

Description

technical field [0001] The invention belongs to the technical field of biology, and in particular relates to a fermentation preparation method of Salmonella pullorum phage lyase. Background technique [0002] At present, pullorum is one of the 14 kinds of animal diseases that the Ministry of Agriculture of my country requires to focus on monitoring. The harm of its pathogenic bacteria to poultry seriously restricts the development of poultry industry. Purification of pullorum in chicken farms is the best way to eradicate the disease. Antibiotics are traditional drugs for the prevention and treatment of pullorum, but due to the long-term widespread use and even abuse of antibiotics, drug resistance is becoming more and more serious, making the prevention and removal of pullorum more difficult. As new antimicrobial agents, phages and their lytic enzymes have been widely studied and applied in improving food and medical safety. Therefore, it is of great significance to screen ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N9/88C12N7/00C12N15/10C12N15/60C12N15/81C12N1/19C12R1/84
CPCC12N9/88C12N7/00C12N15/1003C12N15/815C12N2795/00022
Inventor 冯新宋战昀
Owner JILIN UNIV
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